Difference between revisions of "Part:BBa K4886002"
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<div class="unterschrift"><b>Figure 4 Growth comparison of Ct(Pthl F/Xpk-BD) and Ct(Pthl F/Xpk-QS)</b> | <div class="unterschrift"><b>Figure 4 Growth comparison of Ct(Pthl F/Xpk-BD) and Ct(Pthl F/Xpk-QS)</b> | ||
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<div class="unterschrift"><b>Figure 4 Levels of acetyl phosphate in Ct(Pthl F/Xpk-BD) and Ct(Pthl F/Xpk-QS)</b> | <div class="unterschrift"><b>Figure 4 Levels of acetyl phosphate in Ct(Pthl F/Xpk-BD) and Ct(Pthl F/Xpk-QS)</b> | ||
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Revision as of 16:12, 11 October 2023
Pthl-F/Xpk(QS)
It is a part that is responsible for expressing F/Xpk from Bifidobacterium adolescentis ATCC 15703 with Pthl promotor. It consists of Pthl sequence (BBa_K3443002), strong ribosomal binding site (RBS) sequence (BBa_K103015), F/Xpk sequence (BBa_K4886000) and terminator sequence (BBa_K3585002). F/Xpk is a gene that encodes phosphoketolase. Phosphoketolase is an enzyme with both the Fpk and Xpk activity. It catalyzes the conversion of fructose-6-phosphate (F6P) to erythrose-4-phosphate (E4P) and acetyl-phosphate (AcP), and the conversion of xylulose-5-phosphate (X5P) to glyceraldehydes-3-phosphate (G3P) and AcP.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal AgeI site found at 808
- 1000COMPATIBLE WITH RFC[1000]
Results
(1)Plasmid construction
We used Pthl-adhE2 from BBa_K4408008 as the template and X-pMTL-F and X-pMTL-R as primers to obtain a X-Pthl vector (5461bp) by amplification. We used the Bifidobacterium adolescentis ATCC 15703 genome as a template to amplify the phosphoketolase gene [FXpk(QS)] fragment (2478bp). The vector and fragment were confirmed by gel electrophoresis (Figure 2 and 3). The FXpk(QS) fragment was ligated to the X-Pthl linear vector, using Gibson assembly. The plasmid was then transformed into E.coli JM109. After colony PCR for the transformed bacterial colonies, positive colonies were inoculated, and plasmids were extracted. The recombinant plasmid pMTL-Pthl-FXpk(QS) obtained was confirmed by gene sequencing.
(2 )Transfection of C. tyrobutyricum and its growth
(3)Product yield of the transfected strain